The ultimate objective of this research project is to develop osteoinductive extracellular matrix constructs for bone regeneration by culturing bone marrow stromal cells on a three-dimensional biodegradable polymer scaffold with a flow perfusion bioreactor. We hypothesize that the gene expression of the bone marrow stromal cells for osteogenic growth factors can be affected by varying the culture parameters. We further hypothesize that the gene expression profile of the seeded cells will be predictive of the osteoinductivity of the resulting in vitro generated extracellular matrix constructs and that the osteoinductivity of the extracellular matrix constructs will be enhanced through demineralization when implanted at an ectopic site. We propose a set of two studies specifically aimed at examining these hypotheses. We will investigate the effects of fluid flow rate and in vitro culture duration upon the gene expression of osteogenic growth factors by rat bone marrow stromal cells, which are seeded in three-dimensional biodegradable polymer scaffolds and cultured in a flow perfusion bioreactor developed in our laboratory. We will evaluate the correlation between the gene expression profile of cells seeded within the scaffold during in vitro culture and the osteoinductivity of the resulting extracellular matrix construct at an ectopic site in a rat model in vivo. We will also determine the effect of the removal of the mineralized component of the extracellular matrix construct upon the ability of the construct to induce new bone formation at an ectopic site. This project will provide clinically valuable information regarding bone tissue-inducing orthopaedic biomaterials. The relevance of the information gathered is becoming increasingly important due to renewed concern for the safety of non-degradable implants and the potential for disease transmission with allografts.